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Title: | 黑腹果蠅長非編碼RNA特性研究 Characterizing Long Non-coding RNAs in Drosophila melanogaster |
Authors: | Mei-Ju Chen 陳玫如 |
Advisor: | 陳倩瑜 |
Co-Advisor: | 李文雄 |
Keyword: | 整合性研究,黑腹果蠅,長非編碼RNA,RNA定序技術,染色體免疫沉澱定序技術, Integrative research,Drosophila melanogaster,Long non-coding RNA,RNA-seq,ChIP-seq, |
Publication Year : | 2016 |
Degree: | 博士 |
Abstract: | 次世代定序技術(Next-generation sequencing; NGS)開啟RNA領域研究的新紀元。過往認為只是轉錄訊號擾動的長非編碼RNA (long non-coding RNA; lncRNA),已由許多研究證實其在許多重要生理機制中扮演要角。然而,現今文獻對於重要模式生物—黑腹果蠅(Drosophila melanogaster)的lncRNA瞭解仍相當有限;究其原因,乃黑腹果蠅lncRNA的基礎資訊之稀缺所致。因此,本論文追根溯源,由四個面向對黑腹果蠅lncRNA進行系統性探究—(1) 收集與發現:本論文開發一生物資訊方法,自我們產生的組織特異性RNA-seq資料鑑定出為數不少的新lncRNAs,並與公開資訊可收集之已知lncRNAs整合,呈現迄今最新之黑腹果蠅lncRNA資料集;(2) 特性註解:本論文採用大量的RNA-seq與ChIP-seq資料集(總計93組)增進現有lncRNA的註解資訊如轉錄方向與染色質特徵之品質,並進而觀察摘要出黑腹果蠅lncRNA的一般特性;(3) 基因表現:本論文以RT-qPCR實驗驗證了挑選之lncRNA的基因表現,並彰顯RNA-seq技術平台用於發現lncRNA的結果具有相當的可信度;(4) 轉錄調控:本論文提出一結合序列特徵探勘之生物資訊方法,系統性分析轉錄因子結合位(Transcription factor binding site; TFBS)於lncRNA啟動子出現與否,以及其與lncRNA基因轉錄調控的關聯性。結果顯示,當使用核小體佔據與跨物種保留性資訊,於共表現之編碼基因集進行序列探勘,其所得的序列特徵(或稱順式因子;cis-element),多數與已知的TFBS相似;此外,這些順式因子可在共表現之編碼基因與lncRNA基因的啟動子區域同時觀察得見(較常見於第三期幼蟲至雄蟲階段共表現群集),顯示出共表現之編碼基因與lncRNA基因具有被共同調控的可能性。簡言之,本論文彰顯系統性整合研究的優點,透過基因體與轉錄體資料的整合,大幅加速鑑別lncRNA的特性;而所得之觀察結果可作為黑腹果蠅lncRNA功能研究的堅實基礎。 Recent advances in sequencing technology have opened a new era in RNA studies. Novel types of RNAs such as long non-coding RNAs (lncRNAs) have been found to play essential roles in biological processes. However, only limited information is available for lncRNAs in Drosophila melanogaster, an important model organism. Thus, this thesis aims at chracterizing fruit fly lncRNAs from four aspects: (1) collection and discovery; (2) annotation; (3) expression; and (4) regulation. I developed a computational approach to discover novel lncRNAs from the newly generated tissue-specific RNA-seq data, and then I combined the discovered lncRNAs with previously published lncRNAs into a curated dataset. Next, numerous RNA-seq and ChIP-seq datasets (93 sets) were used to improve the lncRNA annotation such as transcriptional direction and presence of conventional chromatin signatures. With these efforts, I summerized general characteristics of fruit fly lncRNAs in the thesis. In addition, I used RT-qPCR experiments to validate the expression of some randomly selected lncRNAs and demonstrated that RNA-seq is a reliable platform to discover lncRNAs. Moreover, I proposed a method to incorporate de novo motif discoveries to systemically investigate the presence of TFBSs in lncRNA promoters and how it is related to the regulation of lncRNA expression. The result revealed that most of the motifs (cis-elements) discovered from the co-expressed coding gene promoters are similar to the annotated TFBSs, where the motif dicscovery procedure considerd the information of nucleosome occupancy and evolutionary conservation. I also found that common cis-elements were usually observed in the promoters of the co-expressed coding and lncRNA genes in the development stages from L3 to male adlut. In conclusion, this thesis demostrated that integration of genomic and transcriptomic data can largely facilitate lncRNA discovery and characterization, and provided a solid foundation for studying the functions of lncRNAs in D. melanogaster. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/3844 |
DOI: | 10.6342/NTU201601384 |
Fulltext Rights: | 同意授權(全球公開) |
Appears in Collections: | 基因體與系統生物學學位學程 |
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